Our project will provide an integrated study of locus-specific genetic changes that will be measured both in vitro following a genotoxic challenge and in a variety of in vivo tissues. Specifically, in a series of 300 patients with non-small-cell lung cancer undergoing surgery, we propose to assess in vitro tobacco mutagen-induced genetic damage in peripheral blood lymphocytes and to correlate these findings with similar genetic changes, presumably caused by tobacco exposure, in cells obtained from bronchial brushings and surgical specimens of adjacent normal and malignant lung cancer tissues. We hypothesize that there will be concordance in the severity of these site-specific chromosomal lesions in the target, intermediate, and surrogate tissues, the extent of which will be quantitated by fluorescence in situ hybridization (FISH). We have selected four specific chromosomal loci to study (two at 3p21.3, one at 10q23, and one at 5p13.2) that appear by pilot data to be important biomarkers for early detection of lung cancer and for which we have identified and developed probes. The first probe harbors the novel candidate 3p21.3 tumor suppressor gene (TSG) GC20, which is a homolog of the translation factor SUl1 gene. The second PAC clone probe (3p21.3 probe #2) harbors multiple putative 3p21.3 TSGs, including 101F6, NPRL2, RASSF1A, FUS1, SEMA3B, and SEMA3F in the specific 3p21.3 600 kb homozygous deletion region, which has been completely characterized and sequenced and is being studied as part of SPORE Project 1. The third probe harbors the pulmonary-associated protein A1 gene (SFTPA1) at 10q23. The fourth probe harbors the human telomerase reverse transcriptase (hTERT) gene at the 5p13.2 amplicon. Loss of the GC20, 3p21.3 probe #2, and SFTPA1 genomic copy numbers is involved in the initiation of lung cancer cell transformation, while upregulation of hTERT is an early event in the progression of lung tumorigenesis. We have also developed a novel assay to determine telomere length in peripheral lymphocytes (quantitative fluorescent in situ hybridization laser scanning cytometry (Q-FISH (LSC)) and predict that telomere length will be comparable in peripheral blood lymphocytes as well as in carcinogen-exposed target lung tissue. Thus, the overall goals of this project are to determine whether the lymphocyte markers that we have already identified as significant predictors of lung cancer risk are an adequate reflection of similar genetic events in the cigarette-exposed target organ tissues. This finding will have substantial implications for future large-scale, population-based molecular epidemiology studies and for early detection of lung cancer. If the panel of cytogenetic biomarkers in lymphocytes is shown to be expressed similarly in target tissue, these markers could be easily and noninvasively applicable in molecular epidemiologic studies and in early detection of lung carcinogenesis.